Based on the administration convenience, transmucosal buccal drug delivery allows special strength points over peroral routes for systemic delivery. It could achieve local or systemic effect and boost drugs’ bioavailability for agents with first pass metabolism. The current study aimed to manufacture and optimize a lavender oil–based nanoemulsion loaded with zaleplon and incorporate it into fast-disintegrating tablets to promote its dissolution and oral bioavailability via oral mucosa. Zaleplon-loaded nanoemulsions were devised with various levels of lavender oil (10% to 25%), the surfactant Sorbeth-20 (35% to 65%), and the co-surfactant HCO-60 (20% to 40%); the extreme vertices mixture statistical design was adopted. The droplet size and drug-loading efficiency were the evaluated. The optimal formulation was transformed into self-nanoemulsified lyophilized tablets (ZP-LV-SNELTs), which were tested for their uniformity of content, friability, and disintegration time with in-vitro release. Finally, the pharmacokinetic parameters of the ZP-LV-SNELTs were determined and compared with those of marketed formulations. The optimal nanoemulsion had a droplet size of 87 nm and drug-loading capacity of 185 mg/mL. ZP-LV-SNELTs exhibited acceptable friability and weight uniformity and a short disintegration time. The in-vitro release of ZP-LV-SNELTs was 17 times faster than that of the marketed tablet. Moreover, the optimal ZP-LV-SNELTs increased the bioavailability of zaleplon in rabbits by 1.6-fold compared with the commercial tablets. Hence, this investigation revealed that ZP-LV-SNELTs delivered zaleplon with enhanced solubility, a fast release, and boosted bioavailability thru oral mucosa which provided a favorable route for drug administration which is suggested to be clinically investigated in future studies It is primarily recommended for the short-term management of insomnia , preferentially at the α1β2γ2 subunit binding site, which is the benzodiazepine binding site in the central nervous system. Thus, it is also used as a strong anticonvulsant medication in pentylenetetrazole and electroshock-induced convulsions , also known as submicron emulsions, miniemulsions, and ultrafine emulsions, have become increasingly attractive options in terms of dosage form designs and pharmacotherapies. These nanoscale entities serve primarily as vehicles for the delivery of therapeutics, in particular, poorly soluble drugs that are prone to hydrolysis , and silica fume (0.007 mm) were purchased from Sigma-Aldrich . Lavender oil (LV) was obtained from Acros Organics . D-Mannitol was attained from Winlab, Ltd. . Propylene glycol, Cremophor EL, and sodium carboxymethyl were obtained from Spectrum Chemical Manufacturing Corporation . Polacrillin potassium (KYRON T-314) was obtained as a gift from Glenmark Generics, Ltd. . Polyethylene glycol-40, −50, and −60 hydrogenated castor oil , polyethylene glycol-25-stearate (MYS-25V), polyoxyethylene lanolin (Sorbeth-20), polyoxyethylene-C21-ethers (Laureth-21), and sefsol were purchased from Nikko Chemicals Co., Ltd. . A Marketed zaleplon tablet (Sonata) was acquired from Pfizer Ltd. (UK). All other chemicals and reagents used were of analytical grade.2.2.2.2.1.The solubility of ZP in various co-surfactants and surfactants was determined in hopes of finding a variety of self-emulsifying regions that could aid in the development of formulations containing ZP and LV. By dispersing an additional quantity of ZP in 5 mL of each solution individually, the solubility of ZP in several surfactants  and co-surfactants  was determined. Then, the formulations were placed for 72 h in a water bath at 25 ± 2 °C . The mixtures were centrifuged  at 4500 rpm for 15 min after equilibrium of the mixtures was obtained. The supernatants were diluted with methanol. The drug concentration was 240 nm λmax utilizing a previously reported high-performance liquid chromatography (HPLC) method , which had been chosen based on solubility. When the mixture of the ZP, LV, co-surfactant, and surfactant was stirred gently, a transparent nanosized globule emulsion was produced. Therefore, the diagram was thought to be valuable for determining the various amounts of each component of the SNEDDS.The produced mixtures were evaluated for droplet size uniformity and distribution using the Zetatrac instrument . The NE regions could be recognized in the phase diagram when the droplet size was smaller than 1 μm. The NE formulations to be loaded with ZP were then chosen.2.2.3.1) and highest drug-loading capacity (Y2), combinations with various percentages of the three independent variables were utilized. The responses were allocated, and the formulations were optimized using the design of experiment. A mixture statistical design was proposed for the production and optimization of the NE formulations due to its accuracy and effectiveness .2.2.5.Under continuous stirring, NE samples (0.1 mL) were diluted using double-distilled deionized water (10 mL). The dynamic light-scattering approach was utilized to assess the droplet size and polydispersity index (PDI) of the developed formulations   was determined using the following equation:The ZP-loading capacity of the NE formulations was assessed by placing certain known quantities of ZP in vials containing 1 mL of each plain NE formulation. The vials were kept for 24 h in a water bath at a temperature of 25 ± 2 °C until equilibrium was reached. After reaching equilibrium, the mixtures were centrifuged at 4500 rpm for approximately 15 min. To evaluate the loading capacity for each SNEDDS combination, the supernatants were collected and diluted with methanol, and the ZP concentration was measured at a λ1) and drug-loading capacity (Y2) of various ZP-LV-SNEDDS by mixture design are depicted in Components and the observed mean globule size  of the determined responses’ models and for choosing the model that best fit the gathered data. A p-value of less than .05 was considered significant for the models. The components of the optimized ZP-LV-SNEDDS formulation were determined. Following optimization, the formulation (50 mL) was produced and assessed for droplet size and drug loading and then formulated into a lyophilized tablet.The statistical program StatGraphics Centurion XV software, version 13.2.05, and a regression equation were utilized to link the variables with the measured responses . The degrees of freedom F-ratio and 2.2.8.The optimized ZP-LV-SNEDDS was formulated into lyophilized tablets using several additives according to a previously published method (43-45). The pharmaceutical excipients utilized in the zaleplon–lavender oil–self-emulsifying lyophilized tablets (ZP-LV-SNELTs) were 2% porous fumed silica and 2% microcrystalline cellulose (Avicel) as adsorbents, 1.5% KYRON T-314 as a superdisintegrant, 0.5% sodium carboxymethyl cellulose, and 2% gelatin, and 1% mannitol was used as a structure-forming excipient. Briefly, a specified amount of the optimized ZP-LV-SNEDDS containing an amount of ZP sufficient for a 50 tablet was mixed with 50 mL of a gelatin solution (2% w/v) with a magnetic stirrer. The remaining substances were added with continous homogenization at 300 rpm until a homogenous dispersion was formed. The formed dispersions were emptied into a pocket blister pack, which was subjected to a lyophilization process to produce ZP-LV-SNELTs containing 10 mg of ZP in each individual tablet. The lyophilized tablets were kept at room temperature in a desiccator for further evaluation.2.2.9.The ZP-LV-SNELTs were evaluated according to the method described in USP 28/NF23 with regard to different factors, including weight uniformity, thickness, visual appearance, and drug content, in order to confirm the uniformity of the manufactured tablets . In the in-vivo investigation, 12 albino male rabbits weighing between 2 and 2.5 kg were employed. The animals were fasted for 12 hours before the start of the experiment, and they were split into two groups of six rabbits each. The commercially available ZP tablet (reference standard) was given to group 1 at a dose of 1 mg/kg, and group 2 received the optimized ZP-LV-SNELTs at the same dose. Blood samples (1 mL) were withdrawn from the rabbits’ ear veins before drug administration and then again after 5, 15, 30, 45, and 60 min, as well as after 1.5, 2, 3, 4.5, 6, 9, 12, and 24 h. The samples were kept at 20 °C for further analysis.2.2.10.2.max), time to peak drug concentration (Tmax), half-life (t1/2), and mean residence time (MRT). The linear trapezoidal rule was used to determine the area under the curve (AUC0–24). The relative bioavailability and the AUC0–∞ of the optimized ZP-LV-SNELTs were also calculated. Pharmacokinetic parameters of the optimized preparation and the reference tablet were statistically compared using the one-way ANOVA at a significance level of p-value less than 0.05 using an SPSS statistical program.A plasma concentration-time curve was constructed, and the data were fed into the WinNonlin Nonlinear Estimation Program to determine the maximum serum concentration , Sorbeth-20 (B), and HCO-60 (C) and their effects on the droplet size of the ZP-LV-SNEDDS. The chosen model had an adjusted Rp-value of less than .0001. Interestingly, parameter A  had the greatest effect on the droplet size because it gained the highest coefficient (170) when compared with Sorbeth-20 (55.61) and HCO-60 (55.62). Further, the interaction terms of parameter (A) attained higher coefficient values compared with the interaction terms that did not include factor (A).It was observed that all of the investigated variables had a significant effect on the droplet size of the NEs at a The increase in droplet size of the ZP-LV-SNEDDS with the increase in percentage of LV might be ascribed to the expected decrease in amounts of surfactant and co-surfactant used, which would decrease the ability of surfactant and co-surfactant to lower the droplet size and lead to the formation of larger droplets. Comparable outcomes were previously reported , Sorbeth-20 (B), and HCO-60 (C) on the ZP-LV-SNEDDS’ capacity to contain the drug. The model had an adjusted Rp-value of less than .0001.As can be seen from the equation, all of the examined variables had a significant effect on the drug loading at a However, the percentage of LV had the greatest effect on the drug loading; it acquired the highest coefficient (660.20) when compared with Sorbeth-20 (50.84) and HCO-60 (39.06). Additionally, the interaction terms containing factor (A) had higher coefficient values than interaction terms that did not contain factor (A).The increase in drug-loading capacity due to the increase in percentage of LV could be due to the high affinity of the hydrophobic drug ZP for the organic phase; hence, the higher the percentage of oil, the more space available to accommodate the drug. It can also be seen from the figure that increasing the percentage of Sorbeth-20 decreased the drug-loading capacity of the formulations. This could be ascribed to the amphiphilic nature of factor B, which contributed to its ability to decrease the interfacial tension between the aqueous and organic phases, which might lead to the escape of the loaded drug into the surrounding aqueous phase similar reports were found in literature , and this confirmed the precision and validity of the chosen models.By examining the data, an optimal NE formulation was manufactured with the most suitable specifications. The utilized software offered diverse solutions using many combinations of amounts of the variables. The optimum formulation consisted of 13.3% LV, 49.0% Sorbet-20, and 37.7% HCO-60. The optimal formulation had a droplet size of 84.48 nm and a drug-loading capacity of 180.28 mg/mL, with a desirability of 0.812. 3.7.2 values of the dependent variables matched well, and this indicated the ability of the design to predict the capacity. More importantly, the actual and expected ratios recorded for the experimental and predicted responses had a low percentage of error and good residuals; this indicated the lack of curvature in the responses and the validity of the model, as shown in As previously stated, the predicted and adjusted R3.8.The ZP-LV-SNELTs were analyzed to determine different parameters. The weight uniformity of the SNELTs met the pharmacopeia requirement, and the results were 150 ± 2.01 mg. The thickness of the SNELTs was 4.982 ± 0.14. The actual drug content was 9.6 ± 0.24 mg, which is equivalent to 96% ± 2 of ZP content when compared with the theoretical drug content . These findings complied with the pharmacopeia limits, which state that the drug content of developed tablets should range from 90.0% to 110.0%. The friability of the tablets was less than 1%, which agreed with the pharmacopeia specifications and indicated good mechanical strength. Regarding the disintegration time, the optimized ZP-LV-SNELTs formulation disintegrated within 30 sec, and this indicated the porous nature of the formulation, which enhanced its dissolution. Comparable results were found in the literature  after 13 min. The enhancement in ZP release from the developed optimal formulations signified the improvement of ZP dissolution in the nanosized drug delivery system, which offered a large surface area for drug dissolution and release  and the time required to reach it (Tmax) were obtained directly from the plasma concentration-time curve (0→t) was estimated by the trapezoidal rule. It was found that the Cmax obtained following ZP-LV-SNELTs administration (18.22 ± 2.35) was remarkably higher than that of the commercial ZP tablet (9.98 ± 1.22) at a p-value of less than .05. Additionally, the Tmax obtained following the optimal ZP-VL-SNELTs administration (0.5 ± 0.25) was considerably lower than that obtained following commercial product administration (1.5 ± 0.25) at a p-value of less than .0002. The t1/2 for both formulations varied nonsignificantly (1.54 for the test formulation and 1.60 for the standard formulation). The area under the concentration-time curve value, which clarifies the extent of drug absorption, was 835.6 ± 41.33 ng h/mL following administration of the optimal formulation. This was significantly greater than that of the commercial tablets (511.7 ± 32.11 ng h/mL) at a p-value of less than .001). Moreover, there was a nonsignificant difference between the MRTs of both the test and standard formulations.The maximum plasma concentration of ZP (Cme curve . The aremax and AUC and lower Tmax obtained after optimal ZP-LV-SNELTs administration could be ascribed to the fast dissolution of that formulation and the greater absorption of drug from it. This may be due to the applied nanosized drug delivery system, which offers a large area for release and better absorption. Similar results were reported in the literature (Khalifa et al., The greater C4.The developed NE had a high capacity for enhancing the solubility and dissolution of ZP. A pseudoternary phase diagram was constructed to obtain the best levels of LV, Sorbeth-20, and HCO-60 to allocate the most satisfactory NE region for the formulations. The optimal NE had a droplet size of 87 nm with reasonable homogeneity and a drug-loading capacity of 185 mg/mL. The developed ZP-LV-SNELTs had acceptable friability, weight uniformity, and a fast disintegration time, in addition to a 17-fold faster in-vitro release rate, compared with the commercial ZP tablets. Further, the optimized ZP-LV-SNELTs had an increase of approximately 1.6 in drug bioavailability in rabbits compared with the marketed product. In summary, this study highlighted the ability of LV-based self-nanoemulsified tablets to provide better drug solubility and dissolution and, thus, a faster onset of action with better bioavailability. In addition to the high capacity of the oral mucosal delivery to achieve high drug bioavailability thru avoiding first pass metabolism. We recommend studying the prepared formulation clinically in humans as a future prospect of the study.


Caenorhabditis elegans, many genes involved in the formation of the cuticle are also known to influence body size and shape. We assessed post-embryonic growth of both long and shortC. elegans body size mutants from the L1 to L4 stage. We found similar developmental trajectories of N2 andlon-3animals. By contrast, we observed overall decreases in body length and increases in body width of testeddpymutants compared to N2, consistent with the Dpy phenotype. We further show that the dynamics of animal shape in the mutant strains are consistent with a previously proposed “Stretcher” growth model.In Caenorhabditis elegans, a free-living nematode, presents a versatile genetic model system to study how the processes of growth and development are regulated.C. elegansmatures to an adult after multiple molting events during which time animals synthesize a new exoskeleton (cuticle) and expel their old one. This cuticle is a complex, multi-layered structure primarily composed of collagens. As animals progress through their life-cycle, the structure and thickness of the cuticle changes but its role in the maintenance of body morphology and integrity remains. To date, 21 cuticle collagen mutants have been identified that cause a range of body morphology defects (Page and Johnstone 2007). Some of these mutants exhibit a disproportionate reduction in body size compared to the wild type, and others are noticeably larger than the wild type , clearly demonstrating the importance of the physical structure of the cuticle on growth. Analyzing the characteristics of size during development in variousC. elegansbody shape mutants is central to understanding the role these genes have on body growth.Body size largely influences a variety of organismal traits, including: growth, reproduction, metabolism, and lifespan. For this reason, the factors that control organism size, particularly during development, have been explored in many systems.C. eleganscuticle mutants. We chose mutants that were both reportedly shorter (dpy-1(e1), dpy-5(e61)) and longer (lon-3(e2175)) than the wild type  . We then collected high-precision measurements of animal fluorescence , length , and width  from the L1 stage into the L4 stage. As we previously demonstrated , by exposing animals to ingestible fluorescent beads, we can use oscillations in fluorescence as a proxy for feeding behavior. To characterize larval progression, periods of decreased feeding are associated with molt events. Doing so, we noticed thatlon-3(e2175) animals follow similar developmental trajectories to the N2 wild-type strain. By contrast, we detected a delay in the molt timing of bothdpy mutant strains, withdpy-1(e1) undergoing each larval transition later than all other tested strains. We also observed a marked decrease in animal length and increase in animal width noticeable after the L1 stage in bothdpystrains, consistent with their characteristic Dpy phenotype. Interestingly, we noted little size divergence betweenlon-3(e2175)and N2 animals during our time course.We performed a high-resolution longitudinal study of growth in a selection ofC. elegans body shape, and found that model-predicted shape changes were consistent with data from the N2 wild-type strain . Given this result, we proposed a “Stretcher” model for growth whereinC. elegans sense changes in cuticle elasticity, in tandem with other regulatory mechanisms, to control growth rate and determine developmental transitions. Given the structural impacts caused by thedpy-1(e1), dpy-5(e61), andlon-3(e2175) mutations, we sought to determine whether the shape dynamics predicted by the Stretcher model would be consistent with the mutant data. By analyzing the relationship between measured animal length and width over time, we were able to detect the linear stretch, nonlinear stretch, and relaxation regimes predicted by the Stretcher model  and observed in our previous work . If the cuticle responds to internal pressure from growth linearly, then the ratio of change in width and length (W/L) is expected to be constant. If the cuticle stretches beyond some threshold, it exhibits nonlinear response and the ratio will change as the cuticle becomes more stiff in either length or width direction. The linear regime indicated by constant slope at the start of the larval stage and nonlinear stretch regimes indicated by an increase in slope at the end of the larval stage were most apparent in the L2 stage of the N2 strain,lon-3(e2175), anddpy-5(e61). We did not observe as significant an increase in slope indpy-1(e1), possibly due to changes in cuticle elasticity for this mutant, or in the other larval stages, possibly due to challenges in measuring smaller L1 animals and population desynchronization in the L3 stage as animals age. Following the non-linear stretch regime, we expect to observe a post-molt relaxation as the old stretch-restricted cuticle is shed and the animal changes shape to fill the new cuticle. Notably, although we observed a post-molt relaxation period in all strains, we found differences in the dpy mutants when compared to the N2 strain and thelon-3(e2175) mutant (Extended Data). Indpy-1(e1)animals we see post-molt increase in length accompanied by a corresponding decrease in width, consistent with dynamics observed in the N2 strain andlon-3(e2175)animals. However, we noted a marked decrease in the magnitude of relaxation. By contrast, indpy-5(e61)animals, we noticed a change in the direction of relaxation with animals appearing to exhibit post-molt relaxation in width as opposed to length. This difference in relaxation could be a result of changes in cuticle stiffness caused by thedpy-5(e61) mutation, as previous work has revealed thatdpy-5(e61)animals are less stiff than N2 animals .Measurements of both animal length and width allowed us to assess changes in body shape in addition to size. Motivated by changes in the body aspect ratio of animals that we observed at larval-stage transitions, we previously modeled a physical mechanism by which constraints on cuticle stretch could cause changes inC. elegans larval growth. We found that thedpy-1(e1)ordpy-5(e61) mutations strongly impacted organism size and developmental speed. Although it is unclear from our work how much each mutant could interfere with signaling processes associated with molt timing, it is evident that cuticle properties strongly influence growth dynamics at larval-stage transitions.This study investigated the influence of cuticle mutations onWorm cultureC. elegansNatural Diversity Resource . All other strains were provided by the CGC, which is funded by NIH Office of Research Infrastructure Programs (P40 OD010440). Animals were cultured at 20°C on 6 cm plates of modified nematode growth media (NGMA), which contained 1% agar and 0.7% agarose seeded withE. coliOP50 bacteria .The laboratory strain N2 was obtained from theHigh-throughput growth assayMeasurements of body size and fluorescence were measured as previously described . Briefly, strains were propagated for three generations, bleach-synchronized, and titered at a concentration of 1 embryo per µL into 250 mL flasks for a total of 250,000 embryos. The following day, arrested L1s were fed HB101 food at a final concentration of OD20 in a final flask volume of 100 mL K medium and HB101 food. Animals were grown with constant shaking at 20°C. Flasks were sampled each hour beginning one hour after feeding and continuing for 42 consecutive hours. At each hour, 800 µL was removed from each flask and incubated with fluorescent polychromatic beads  for 10 minutes with shaking. Following the bead incubation, animals were transferred to a 96-well microtiter plate, treated with sodium azide, imaged with an ImageXpress Nano , and scored using a large-particle flow cytometer . COPAS BIOSORT was used to collect measurements of animal length (TOF), optical extinction (EXT), and red fluorescence for every animal in each well.Data processingCOPAS BIOSORT data were processed as previously described . To remove non-animal objects such as bacterial clumps, shed cuticles, and next-generation larval animals from the time-course data. Data for each well were summarized, generating median well measurements. TOF and norm.EXT data were then converted to microns. This conversion was performed by first collecting manual measurements of animal size from images using the free Java image-processing program ImageJ . Briefly, length was measured from head to tail, and width was measured at the widest point of the animal. Five animals were measured per well across thirty total wells for each hour. Pixels were converted to µm using a conversion factor of 3.2937 pixels/µm. Only the unit-corrected data were used for further analysis.lon-3: 16-22,dpy-1: 19-26, anddpy-5: 19-24. Lastly, the L3 hours were: N2: 27-31,lon-3: 27:32,dpy-1: 31-35, anddpy-5: 30-34. The L4 stage was excluded because of high variability in the population. We next applied a local kernel regression to smooth the population measurements and calculated mean and standard deviation using bootstrapping. The ratio of the change in width to length over time, calculated from the local slope of the population data, was calculated to determine cuticle properties across larval stages. All files and code for analysis and generation of figures and tables are archived on GitHub (https://github.com/AndersenLab/cuticle-mutants-micropub).“Stretcher” model analysis of shape dynamics was performed as previously described . Briefly, length and width data were extracted from the full population data by selecting time periods that were in active growth and omitting time periods where post-molt relaxation could be observed. The hours defining these growth periods for the L1 stage were, 1-10. For the L2 stage the hours were: N2 and10.22002/d60qd-yy629Description: Median length (x-axis) is plotted against median length (y-axis) to visualize changes in body aspect ratio over time. Each point corresponds to the median length or median width of animals at each experimental hour. Post-molt relaxation is observed in WT, lon-3(e2175), and dpy-1(e1) as a slight decrease in median animal width. Post-molt relaxation is observed in dpy-5(e61) as a slight decrease in animal length. . 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Bacillus sp. strain TM2, an aerobic Gram-positive bacterium, was isolated from a solution of ground Tetragnatha maxillosa spider in saline solution in Japan. Here, we report the complete genome sequence of this bacterium, which has a 3.67-Mbp genome containing 3,702 protein-coding sequences, 24 rRNA-coding sequences, and 81 tRNA-coding sequences. Bacillus sp. strain TM2, which was isolated from a solution of ground Tetragnatha maxillosa spider isolated in Japan . The method for isolating strain TM2 from the spider was the same as that reported previously  according to the manufacturer’s instructions, and the library's quality was confirmed using a Fragment Analyzer system and the double-stranded DNA (dsDNA) 915 regent kit . The reaction time for enzyme cleavage was 4 min, and an average fragment length of 442 bp was prepared. Circularized DNA was prepared using the prepared library and the MGIEasy circularization kit according to the manual. DNBSEQ 2 × 200-bp paired-end sequencing was performed using the DNBSEQ-G400 sequencer according to the manufacturer’s instructions. The adapter sequences were removed using Cutadapt (v2.7)   was usedHigh-quality short-read and long-read sequence data were assembled under the default conditions of Unicycler (v0.4.7) , and theBacillus sp. strain TM2 is AP025262, and that for the extrachromosomal plasmid is AP025263. Raw sequencing data were deposited in the SRA under accession number DRA013015.The GenBank accession number for the whole-genome sequence of


Spiraea hypericifolia L. is affiliated with the section Chamaedryon Ser. of the genus Spiraea L. (Rosaceae). Similar to many other Spiraea species, S. hypericifolia most often accumulates flavonols among other flavonoids, in particular quercetin and its derivatives. An ethanol–water extract from the aerial part of S. hypericifolia collected in the vicinity of the Ilyichovo settlement  was analyzed by liquid chromatography with high-resolution mass spectrometry. Primary and secondary metabolites were found in the extract; structural interpretation consistent with quercetin and its derivatives was proposed for 10 of them. Major compounds were various glycosides of quercetin containing glucose (four compounds), galactose (one compound), xylose (two compounds), arabinose (one compound), or rutinose (one compound) as a carbohydrate residue. Isorhamnetin and 3-O-methylquercetin-3′-O-β-D-glucopyranoside were identified among methyl-containing compounds. The latter compound and reynoutrin, rhamnetin-3-O-β-D-xylopyranosyl-β-D-glucopyranoside, and quercetin-3-O--β-D-glucoside have not been previously found in S. hypericifolia. Data on the presence of quercetin and its derivatives in the extract of S. hypericifolia expand the understanding of the possible practical use of this plant. In addition, the microscopic features of S. hypericifolia leaves were studied. The diagnostic features of the leaf blade necessary for the authentication of raw materials were revealed: straight-walled epidermis cells, stomata located on both sides of the leaf blade (amphistomatic type), two types of trichomes, and wrinkled cuticula with nodi. The main anatomical diagnostic features of the leaves of S. hypericifolia were determined, which makes it possible to assess the authenticity of the raw material. Flavonoids are a large family of natural compounds that are characterized by various structures, high and diverse biological activities, and low toxicity . The devQuercetin, that is, 3,5,7,3′,4′-pentahydroxyflavone, is a flavonoid that belongs to the flavonol subclass and is of particular interest because it can be found in most of the studied plants . QuercetQuercetin is a pharmacologically active compound because it has various biological effects. In most cases, researchers focus on its antioxidant potential ,6,7,8; hSpiraea L. species are flavonoid-accumulating plants whose extracts may contain up to 120 mg/g flavonoids [Spiraea genus, E.A. Karpova and N.P. Lapteva concluded that this genus’s flavonoid compounds are mainly represented by flavonols, with quercetin accounting for 50% to 90% of aglycons, and the remaining flavonoid compounds are represented by kaempferol and isorhamnetin and their derivatives [Spiraea species also showed that the majority (over 25) of flavonoid compounds are quercetin and its derivatives [Spiraea species, we attempted to study phenolic components of Spiraea representatives from Asian Russia, including Spiraea hypericifolia L. [S. hypericifolia L. are fragmentary; the plant has been found to contain such compounds as flavonols, proanthocyanidins, and catechins [S. hypericifolia have not been studied separately.avonoids . From daivatives . A literity over  of flavofolia L. ,27. Dataatechins . QuercetSpiraea hypericifolia L. is affiliated with the section Chamaedryon Ser. of the genus Spiraea L. (Rosaceae). This plant is a shrub up to 80 (150) cm in height with brown smooth limbs (puberulent in young plants). It has long-ellipsoidal or lanceolar, grayish-green, smooth-margin leaves. Flowers are white and are bound in an attached umbrella blossom cluster; the fruit is a leaflet. S. hypericifolia occurs in Europe, the Caucasus, Central Asia, Russia, Mongolia, and China [nd China ,30. It gnd China .Spiraea humilis Pojark. and Spiraea salicifolia L., from Russia has revealed diagnostic features that can be employed for interspecies diagnosis for their practical medicinal use [Spiraea section can be diagnosed only according to the structure of the core and several quantitative features. Leaves of the species under study are diagnosed on the basis of thickness of the lamina near the midrib and in the areas distant from ribs, the thickness of the paxillate mesophyll, and the presence or absence of trichomes [S. hypericifolia leaf has not been studied in Russia.The determination of the anatomical features of medicinal plants helps with quality control of the production of phytotherapeutics . Anatomiinal use . It is rrichomes . The anaS. hypericifolia extracts for quercetin and its derivatives and (ii) anatomical and diagnostic leaf analysis.This study is aimed at (i) assaying The analysis of LC–HRMS data was performed to characterize primary and secondary metabolites, of which 10 were structurally interpreted as quercetin and its derivatives .S. hypericifolia, only isorhamnetin is a methyl-containing derivative of quercetin, with the most substances detected being various quercetin glycosides (O-methylquercetin-3′-O-β-D-glucopyranoside contains both a methyl group (position C-3) and a carbohydrate residue (position C-3′). The carbohydrate part of the molecule is composed of mono- and disaccharides. Monosaccharides include pentoses (arabinose and xylose) and hexoses , whereas disaccharides include rutinose. The carbohydrate moiety is usually attached at the C-3 position. In some cases -β-D-glucoside), glucoside is additionally acylated with malonic acid on the sugar’s hydroxyl group.Among substances identified in the water–ethanol extract of ycosides . 3-O-metS. hypericifolia but also in other Spiraea species has been proved by other researchers. Literature data on the phytochemical content of certain species vary (data on some taxa are scarcer than fragmentary because they are reported in one article only); therefore, it is a foregone conclusion that the presence of the flavonoid family in Spiraea is diverse. As shown in Spiraea species contain flavonols. We suppose that in terms of the general diversity of flavonols contained in Spiraea, attention should be drawn to S. salicifolia. Literature data on this species are more or less representative (34 flavonols) because this taxon has been a subject of phytochemical studies more often than others. It is difficult to assess the diversity of flavonols in Spiraea by means of studies on such phytochemically underinvestigated species as S. albiflora, S. brahuica, or S. nipponica, in which only one flavonol has been found.A hypothesis that quercetin and its derivatives tend to accumulate not only in S. hypericifolia, whereas quercetin-3-O--β-D-glucoside, reynoutrin, rhamnetin-3-O-β-D-xylopyranosyl-β-D-glucopyranoside, and 3-O-methylquercetin-3′-O-β-D-glucopyranoside have not been found previously in either this species or other Spiraea taxa.Other researchers have reported that rutin, quercetin, avicularin, hyperoside, and isoquercitrin (identified by us) are present in Spiraea species, quercetin has proved to be the most common substance (in 21 species) in various studies; hyperoside has been detected in 14 species; isoquercitrin has been identified in 10 Spiraea species; and avicularin, rutin, and isorhamnetin have been found in 10 species by various authors [Regarding the set of flavonoid compounds in  authors .Quercetin’s pharmaceutical characteristics have been investigated the best to date. Most of the studies deal with its aglycone form; however, plasma analysis performed after administration of quercetin shows that quercetin glycosides are the main circulating compounds . VariousQuercetin glycosides arise via attachment of a sugar to quercetin by replacement of one of the hydroxyl groups, with subsequent formation of a glycoside linkage. Quercetin glycosylation may theoretically take place on any hydroxyl group, whereas most common quercetin glycosides have a sugar group at the C-3 position. The isoquercetin structure has been found to contain glucose attached to the OH group of quercetin at the C-3 position. The addition of galactose to the quercetin molecule at the same C-3 position leads to hyperoside, and the addition of a rhamnosyl group initiates the synthesis of quercitrin. Several quercetin derivatives contain disaccharides, such as rutinose or arabinofuranose; their attachment at position C-3 causes the formation of important compounds: rutin and avicularin, respectively .O-rutinoside (narcissin) and other compounds.There are also some methylated derivatives of quercetin. For instance, rhamnazin contains two methyl groups at positions C-7 and C-3′. Isorhamnetin is another methylated flavonol (C-3′ position), which can be glycosylated, giving rise to quercetin 3-A carbohydrate molecule attached to quercetin aglycone has been shown to improve water solubility, absorption, and other properties . The exaMoringa oleifera Lam. (Moringaceae) [Morus alba L. (Moraceae) [Apocynum venetum L. [A. hendersonii Hook. f. (Apocynaceae) [O-β-D-xylopyranosyl-β-D-glucopyranoside and 3-O-methylquercetin-3′-O-β-D-glucopyranoside are yet to be researched.Rutin’s pharmaceutical characteristics are studied best among all quercetin glycosides: this compound promotes mammalian smooth muscle relaxation , and rutngaceae) , Morus aoraceae) , Apocynunetum L. , and A. ynaceae) . The theS. hypericifolia leaves were investigated next. Visual examination revealed straight-walled epidermis cells and anomocytic stomata located on both sides of the leaf, being numerous under the leaf (amphistomatic type)  . Two typS. hypericifolia in the literature. S. Kuzieva et al. [S. hypericifolia growing in Uzbekistan. As a result, it has been shown that the leaves of this plant are amphistomatic; the epidermis consists of one row of cells with a thick-walled cuticle layer, and the stomata are not submerged. Leaf trichomes are also described, and according to these data, they are all of the same type: simple, opaque, awl-pointed, very rarely located along large veins. The discrepancy between the literature data and our findings may be explained by differences in the methods for obtaining micropreparations because those authors did not conduct special analyses of trichomes and merely recorded their presence on transverse sections of leaves near large veins. Additional research on this issue is required. Furthermore, for a comparative analysis in the future, it is important to study the anatomical and diagnostic features of other species of Spiraea, especially those with morphological features similar to those of S. hypericifolia.There is not much information about the anatomical features of the leaves of a et al.  researchS. hypericifolia were collected during the flowering period in the vicinity of the Ilyichovo settlement, Shushensky District, Krasnoyarsk Krai (Russia). Voucher specimens  were deposited in the plant material storage room in the Laboratory of Phytochemistry, CSBG SB RAS . Air-dried plant material was mechanically ground to obtain a homogeneous powder with a particle size of 2–3 mm. The dry extract was prepared as follows: The plant material was extracted in a water bath in three replicates (100 mL in the first replicate and 75 mL in the second and third replicates) with 70% ethyl alcohol for 8 h at 60 °C. After cooling, the combined filtrates were concentrated in a rotary evaporator to remove the solvent, and then the thick extract was dried in a vacuum drying cabinet to 5% residual moisture. To identify flavonoids, a stock solution of the crude extract was prepared by dissolving the dry extract in 70% ethanol at a 1:1000 ratio.Aerial parts of LC–HRMS was carried out at the Core Facility of Mass Spectrometric Analysis at the Institute of Chemical Biology and Fundamental Medicine SB RAS .S. hypericifolia extract. The chromatographic separation was attained at a 0.4 mL/min flow rate on a Zorbax Eclipse XDB-C18 reversed-phase column  thermostatted at 40 °C. The mobile phase was composed of 0.1% aqueous formic acid (eluent A) and acetonitrile (eluent B). The elution gradient was implemented as follows: from 5% to 70% B for 40 min, followed by an increase to 90% B for 8 min, a decrease to 5% B for 5 min, and re-equilibration under the initial conditions for 7 min.An UltiMate 3000 liquid chromatograph  coupled with a Q Exactive HF mass spectrometer (Thermo Fisher Scientific) was utilized to determine the flavonoid profile of the m/z 200. The following settings of the mass spectrometer were employed: scan range, m/z 80–1200; automatic gain control (AGC), 3e6; injection time, 100 ms; and isolation window, m/z 2.0. The normalized collision energy for the fragmentation of molecular ions was set to 40 eV. Targeted tandem mass spectrometry  was performed in both positive and negative modes at 15,000 FWHM (m/z 200). AGC for dd-MS2 was set to 1e5, with an injection time of 50 ms and a loop count of 5. In the section of dd settings, the AGC target was programmed at 8e3, and the maximum injection time was set to 100 ms. Data were analyzed using Xcalibur 4.0 and the Compound Discoverer 3.1 software (Thermo Fisher Scientific). All the samples, including blank ones, were assayed in triplicate. All the samples were processed in Compound Discoverer 3.1 via a common workflow, “Environmental Unknown ID w Online and Local Database Searches” , MassBank , PlantCyc , and Planta Piloto de Quimica Fina Universidad de Alcala , were chosen in ChemSpider.The parameters set for the electrospray ionization (ESI) source were as follows: electrospray voltage: 3.2 kV in the negative mode and 4.2 kV in the positive mode; capillary temperature: 320 °C; and S lens RF level: 50. Data were obtained by full-scan data-dependent acquisition (FS-dd-MS2) in the positive and negative modes at a resolving power of 45,000 full width at half maximum (FWHM) earches” . A mass https://www.mzcloud.org, last accessed 10 March 2021). If compounds were absent in mzCloud, they were tentatively identified using a ChemSpider search. According to the workflow, the masses extracted from the chromatograms were aligned and filtered to remove (i) background compounds present in the blank sample, (ii) substances that failed to become fragmented, (iii) compound masses that were absent in the databases, and (iv) signals with low intensity.Flavonoids were identified on the basis of both accurate mass and fragment mass “fingerprint” spectra via searches against the spectra of compounds available in the mzCloud database , whereas rutin, avicularin, and hyperoside from Fluka Chemie AG (Switzerland).Leaves from the herbarium were studied visually with the naked eye using a 10× magnifier in accordance with the requirements of the books Herbae and Technique of Microscopic and Microchemical Studies of Herbal Drugs and Herbal Medicinal Products, State Pharmacopoeia of the Russian Federation, XIV edition .Analyzed specimens were clarified by boiling in 5% sodium hydroxide and chloral hydrate. Micropreparations were examined under a BIOSCOP-1 biological microscope with 4×, 10×, and 40× lenses and 7×, 10×, and 15× oculars. Microphotoshooting was performed using a ToupCam FMA050 digital camera (12 megapixels).S. hypericifolia; 10 of these were identified as quercetin and its derivatives. Seven of the identified substances  have pharmacological activities, according to numerous studies. Hence, the therapeutic potential of S. hypericifolia has not been exploited so far. The pharmacological activity of other found substances -β-D-glucoside, rhamnetin-3-O-β-D-xylopyranosyl-β-D-glucopyranoside, and 3-O-methylquercetin-3′-O-β-D-glucopyranoside) requires further investigation. Our data should improve the understanding of the enormous pharmacological potential of S. hypericifolia. Results on the content of quercetin and its derivatives  may be of interest for food and cosmetics industries. The anatomical features of S. hypericifolia leaves were studied too. The anatomic diagnostic features of the raw material can be defined as straight-walled epidermis cells, stomata located on both sides of the leaf blade (amphistomatic type), two types of trichomes, and wrinkled cuticula with nodi. The anatomical features of S. hypericifolia leaves will allow investigators to evaluate the authenticity of raw materials, which have diverse uses.Primary and secondary metabolites were detected in an ethanol–water extract of


Pseudomonas aeruginosa to spread within a hospital critical care setting.To describe the propensity of carbapenem-resistant P. aeruginosa resistant to piperacillin, ceftazidime, cefepime, imipenem, meropenem and ciprofloxacin, isolated from clinical and screening specimens from four critical care units from 2015 to 2020 were analysed. Genotyping was carried out by WGS (Illumina and MinION). MLST, core genome MLST (cgMLST) and resistome analysis was performed and merged with epidemiological data.The study was conducted in a 700-bed tertiary centre in Cologne, Germany. P. aeruginosa isolates were available, of which 20 were carbapenemase producers as follows: blaVIM-1 (n = 1), blaVIM-2 (n = 17), blaVIM-4 (n = 1), and blaNDM-1/blaGES-5 (n = 1). Forty-two of 55 isolates were hospital-acquired. cgMLST revealed three clusters: Cluster 1 ; Cluster 2 ; and Cluster 3 . The blaVIM-2 gene of Cluster 1 was integrated on the chromosome in a class 1 integron (type In59). Using conventional epidemiology, we were only able to confirm two patient-to-patient transmissions and one room-to-patient transmission on three different ICUs within Cluster 1. Isolates from Cluster 2 represented an outbreak occurring in 2019.Fifty-five out of 79 non-duplicate P. aeruginosa. Transmission dynamics differed between carbapenemase- and non-carbapenemase-producing isolates. A continuous acquisition of clonally related ST111 VIM-2 P. aeruginosa, being the main carbapenemase-producing strain, was observed over the whole study period, as well as an overall higher genomic diversity among non-carbapenemase-producing P. aeruginosa.These data give insight into the epidemiology of carbapenem-resistant  Pseudomonas aeruginosa is an environmental bacterium that can colonize the human body. As a leading nosocomial pathogen P. aeruginosa may cause surgical site infections, ventilator-associated pneumonia, catheter-associated urinary tract infections or central-line-associated bloodstream infections in healthcare settings.1 The organism is especially problematic for immunocompromised patients within special units .2 Infections can be difficult to treat because of intrinsic resistance to many antimicrobial agents as well as rapid development of antimicrobial resistance to nearly all available antimicrobials through chromosomal mutations and acquisition of transferable resistance genes.3 Of particular interest is carbapenem resistance mediated by intrinsic resistance mechanisms  or acquisition of a carbapenemase, especially an MBL.3 Carbapenemase production is linked to globally distributed and emerging MDR or even XDR high-risk clones.5 While the emergence of carbapenem-resistant P. aeruginosa is well described, less is known about the different propensity of carbapenemase-producing and carbapenemase-non-producing P. aeruginosa to spread within the hospital setting. This is important, as a relevant part of hospital-acquired infections caused by P. aeruginosa is transmission-associated, either patient-to-patient  or environment-to-patient.7 The hospital environmental is a known reservoir, especially in moist sites. Reports show that contaminated tap water as well as washbasins are linked to transmission events.8–11P. aeruginosa over a 3 year period.12 Using PFGE nearly all VIM-2-producing isolates were clonally related. However, only a few epidemiologically proven transmission events were confirmed, exclusively on several ICUs of the tertiary care centre.12 The present investigation aims to define the local epidemiology and transmission dynamics of MDR/XDR P. aeruginosa irrespective of carbapenemase production in these ICUs of the tertiary care centre over a period of 5 years using a genomics-based approach.A previous study from our research group performed in three hospitals of different levels of healthcare has shown a prevalence of approximately 20% carbapenemase producers, mostly VIM-2, amongst MDR/XDR 13 and previous studies,12 four out of six available critical care units with a frequent detection and/or possible transmission events of carbapenem-resistant and MDR/XDR P. aeruginosa were chosen: three ICUs (ICU 1–3) and one intermediate care unit (ImCU1). ICU1 and ICU3 provide care for surgical patients including burn patients , whereas ICU2 and ImCU1 are primarily reserved for medical patients . Overall, there were approximately 3500 admissions (20 000 patient days) per year on these units. The number of patients colonized/infected with P. aeruginosa was assessed using the laboratory surveillance information system .The study was conducted in a 700-bed tertiary care centre in Cologne, Germany. Based on data of the implemented active surveillance system following the protocol of the German healthcare-associated infection surveillance on intensive care units (ITS-KISS)P. aeruginosa resistant to piperacillin, ceftazidime, cefepime, imipenem, meropenem and ciprofloxacin isolated on the designated wards from clinical and screening specimens from January 2015 to June 2020 were included. This basically corresponds to an MDR/XDR phenotype according to the ECDC/CDC classification.5 Two VIM-2-producing P. aeruginosa isolates detected on other wards of the same hospital analysed in a previous study (PSA-2016-03 and PSA-2017-02) were also included in this study.12All isolates were identified with standard microbiological procedures using the Vitek 2 system  or MALDI-TOF . First susceptibility testing was performed with automated systems  or disc diffusion  and later confirmed by broth microdilution using Micronaut-S Pseudomonas MIC panels  according to the manufacturers’ instructions. MICs were determined for piperacillin, piperacillin/tazobactam, ceftazidime, cefepime, ceftazidime/avibactam, ceftolozane/tazobactam, imipenem, meropenem, aztreonam, gentamicin, tobramycin, amikacin, ciprofloxacin, levofloxacin, fosfomycin and colistin. EUCAST breakpoints  were used for interpretation. De novo assembly was performed using Velvet (version 1.1.04).14 The raw sequencing short reads generated in this project were submitted to the European Nucleotide Archive (https://www.ebi.ac.uk/ena/) under the Project Accession number PRJEB43695.To prepare short-read sequencing libraries, fresh cultures were grown overnight on Mueller–Hinton agar and DNA was isolated using the DNeasy Ultra Clean Microbial Kit  following the manufacturer’s instructions. Sequencing libraries were prepared with the purified DNA using the Ultra II FS DNA Library Prep Kit  for a 250 bp paired-end sequencing run on an Illumina MiSeq sequencer. blaVIM-2 gene, three strains belonging to the same core genome MLST (cgMLST) cluster  were selected for long-read sequencing. DNA was extracted from bacteria grown overnight in Luria broth using the Genomic-Tips 100/G kit and Genomic DNA Buffers kit (Qiagen). Libraries were prepared using the Ligation Sequencing Kit (SQK-LSK109) combined with Native Barcoding Kit (EXP-NBD114) and were loaded onto a R9.4 flow cell  for a MinION sequencing run. Finally, a hybrid assembly of the long- and short-reads was performed using Unicycler.15 The long-read raw data have been deposited to the Sequence Read Archive (https://www.ncbi.nlm.nih.gov/sra) under the BioProject Accession number PRJNA771632.To understand the genetic location of the et al.16 and based on 3867 target genes. During comparison of the allelic profile the ‘pairwise ignoring missing values’ option was turned on. Genomes containing at least 95% of the defined cgMLST targets were included. Isolates with less than 12 different alleles in the cgMLST target gene set were considered as highly related (and termed a cluster). Additionally, based on the assembled genomes, the conventional 7-loci MLST scheme was retrieved from the MLST database.17 Furthermore, acquired resistance genes on assembled genomes were identified by the ResFinder Bacterial Analysis Pipeline v. 2.1.18Relatedness of all isolates was assessed by a cgMLST genotyping approach. Assembled genomes were analysed by SeqSphere+ software  using a validated cgMLST scheme recently proposed by Tönnies blaVIM-2 was annotated and curated manually and visualized using the SnapGene® software  based on the hybrid assemblies. Insertion sequence elements were investigated using the ISfinder database (http://www-is.biotoul.fr).19The genetic environment of P. aeruginosa (single room and use of gowns and gloves). Relevant clinical and epidemiological data was collected from patients’ clinical records or the attending physician. If the collection of the specimen occurred on or before the second day of admission, and there was no prior contact to the healthcare system within the previous 30 days, bacterial isolates were considered community-acquired. If prior contact with the healthcare system  was observed within the previous 30 days and collection occurred on or before the second day of admission, isolates were considered healthcare-associated. If the collection of the specimen occurred after the second day, or if the patient stayed at our hospital within the last 30 days, bacterial isolates were defined as hospital-acquired. Transmission analysis was based on epidemiological data  and genetic data. Proven transmission events were defined as isolation of genetically related isolates (cluster) in two or more patients who were hospitalized during overlapping periods on the same ward  or in the same room with a maximum time interval of 6 months (room-to-patient transmission).12 An interval of 6 months was chosen because transmission of P. aeruginosa from environmental sources can continue over longer periods and can be sporadic.12 Hospital-acquired infections were classified according to the CDC/NHSN definitions.20 Patients without related signs of infection were considered to be colonized.A general rectal admission screening for MDR Gram-negative organisms was in place on all units. Additionally, weekly tracheal secretions were taken from intubated patients, and wound swabs from burn patients (surveillance cultures). Weekly rectal screenings were performed on ICU1 only. Standard and contact precautions were applied for every patient found colonized or infected with MRD/XDR The study was approved by the Ethics Committee of the Faculty of Health of the Witten/Herdecke University (study number S-33/2021).P. aeruginosa clinical and screening isolates were detected from the designated wards during the time period and 55 non-duplicate isolates were available for further analysis.Seventy-nine first MDR/XDR n = 20) the susceptibility rate for aztreonam was 85% and was much lower in non-MBL-producing isolates (n = 34), being 26.4%. All MBL-producing isolates were resistant to ceftolozane/tazobactam and ceftazidime/avibactam, whereas in all non-MBL-producing isolates, susceptibility rates to ceftolozane/tazobactam and ceftazidime/avibactam were both 72.7%. MICs are shown in Table JAC-AMR Online).Susceptibility testing by broth microdilution showed that all bacterial isolates displayed an MDR or XDR phenotype as defined by the inclusion criteria; no isolate was pandrug resistant. One isolate was not cultivable for broth microdilution. The remaining 54 isolates displayed a susceptibility rate of 50%, 61.1% and 100% for tobramycin, amikacin and colistin, respectively. In all MBL-producing isolates , blaVIM-2 (n = 17), blaVIM-4 (n = 1) and blaNDM-1/blaGES-5 (n = 1). Other relevant acquired resistance genes are shown in Table Sixteen of these isolates were previously analysed by conventional genotyping methods.As all patients were from critical care units, devices, surgical and nonsurgical interventions and antibiotic therapy were common Table . The modP. aeruginosa isolates (n = 20) were assigned to five STs, predominantly ST111, but also ST273, ST654, ST235 and ST3618, the latter being a newly assigned ST. A high diversity was detected within the remaining 35 non-carbapenemase-producing P. aeruginosa isolates, which comprised 24 different STs. cgMLST revealed three clusters that were represented by different STs: Cluster 1 , Cluster 2  and Cluster 3 . Within Clusters 1, 2 and 3, a maximum difference of 21, 1 and 5 alleles, respectively, was observed. Carbapenemase production in P. aeruginosa was significantly associated with belonging to a clonal cluster . Table blaVIM-2 gene and the aacA29-like gene. Moreover, all but two isolates from Cluster 1 were hospital-acquired; the other two were healthcare-associated and had contact with two different institutions. Cluster 1 isolates were detected sporadically on all four wards and during the whole study period from 2015 to 2020. Using epidemiological data, we were only able to confirm one room-to-patient and two patient-to-patient transmissions -29a and aacA(6′)-29b, differing in four amino acid substitutions, as well as the integrase gene intI1. Further upstream the antiseptic resistance cassette qacE was detected, followed by sul1 conferring resistance to sulphonamides. The genetic environment of the MBL was highly similar  to the integron In59.23 Finally, the integron was inserted into a transposon TnAs3-like structure   of three ST111 VIM-2-producing isolates from the cgMLST Cluster 1 revealed that the genetic environment of the P. aeruginosa is an important hospital-acquired pathogen causing infections and outbreaks in ICUs.1 Over the last few years we have seen a dramatic increase of MDR isolates worldwide.25 In this study, from a collection of MDR/XDR P. aeruginosa isolates from four critical care units of a tertiary care centre transmission was nearly exclusively observed among ST111 VIM-2-producing isolates over a period of 5 years. Another short-term outbreak caused by ST970 non-carbapenemase-producing P. aeruginosa was observed in vulnerable patients in the burn unit. We observed a higher diversity in non-carbapenemase-producing isolates, most of the isolates being genetically unrelated to each other. Overall, the study reveals a remarkable clonal diversity, with most isolates represented by sporadic single genotypes, and a few epidemic strains.P. aeruginosa infections are disproportionally caused by a small subset of globally distributed ‘high-risk clones’, linked to mutational resistance determinants but also transferable resistance genes.26–28 Traditionally, these clones are classified according to the MLST scheme developed by Curran et al.29 over 15 years ago. Four out of the worldwide top 10 P. aeruginosa high-risk clones  were found in this study.4Studies comparing clonal diversity among susceptible and MDR/XDR resistant isolates have shown a lower diversity among MDR and especially XDR strains and that XDR/MDR 30 Other surveillance studies from the UK and the Netherlands reported that ST111 P. aeruginosa was linked mostly to VIM-2 production.22 Epidemiological data and high resolution genotyping by WGS showed evidence for spread within and between hospitals in different regions of the UK.21 It is important to note that in our study most isolates were hospital-acquired and a limited number of isolates were also present on admission and healthcare-associated. As there is no continuous molecular surveillance on a regional or national level in Germany, we are unable to determine the extent to which ST111 carrying carbapenemases has spread throughout the region. However, VIM-2 is the leading carbapenemase in P. aeruginosa based on data from the German national reference centre.31 Moreover, previous studies have shown ST111 VIM-2-producing P. aeruginosa in Hamburg in 2001,32 and Wendel et al.8 reported the spread of ST111 GIM-1-producing P. aeruginosa in a hospital located close to Cologne. Furthermore, we also demonstrated that the ST111 isolates from the UK and the Netherlands clustered closely, although not overlapping, with those from the current study, highlighting that these high-risk clones are not confined to small geographical regions but have in fact spread to other countries. The genetic environment of the blaVIM-2 gene of the ST111 clone was highly similar to the integron In59 described in a VIM-2-positive P. aeruginosa isolate recovered in 1997 in France and since then in different European countries, and mostly in ST111 isolates.33–35 As high-risk clones tend to harbour several resistance traits they are also linked to aminoglycoside resistance, with the aacA29a gene being the most common determinant in ST111, also confirmed in our study.30In our study the appearance of high-risk clones was also linked to carbapenemase production, mostly ST111 having acquired VIM-2, a combination that has also been described previously.P. aeruginosa is of utmost importance with regard to the clonal structure and mobile genetic elements such as carbapenemases. Both SNP-based and cgMLST-based typing have been successfully applied in various studies.36–39 Recently, several validated cgMLST schemes were published enabling a standardized approach and a consistent nomenclature.41 The cgMLST scheme proposed by Tönnies et al.16 (3867 targets) used in this study was comparable to an ad-hoc cgMLST scheme previously established by one of the authors (4547 targets).39Microbiological and infection control monitoring of carbapenem resistance in P. aeruginosa is either endo- or exogenous and can subsequently lead to transmission chains. Sporadic or low-frequency transmissions from the environment (mostly moist sites) to the patient are difficult to trace back epidemiologically as shown in several studies.11 However, we were unable to confirm many transmission events within the hospital despite sporadic appearance of the clone. A hidden environmental reservoir  or complex epidemiological links might be an explanation. There is growing evidence supporting a water-free patient environment and removing sinks from the patient’s room to eliminate patient-side biofilm reservoirs.43 This is possible on the ICU where the patient generally does not need a bathroom. Nevertheless, we did not find a pattern of room transmissions as patients from the biggest cluster were found on all four wards.Depending on the mode of transmission, different infection and control approaches are needed. Individual nosocomial acquisition of 44 However, carbapenemase production is often linked to MDR/XDR phenotypes.45There are a few limitations in this study. Unfortunately, we were not able to provide full prevalence data, as only two-thirds of the non-duplicate isolates detected during this period were available for further study. Moreover, transmissions by unidentified colonized patients might have been overlooked as there was no periodic rectal screening in place on all units. However, we are confident that the study gives a good overview of the epidemiological pattern. Secondly, the inclusion criteria were probably not sensitive enough to detect carbapenemases, as we chose to include isolates based on the German classification guideline for Gram-negative MDR organisms .in silico using a web-based tool. This is more complex and not completely validated yet and out of the scope of our investigation, which is basically epidemiological.46 Fourthly, we did not conduct environmental sampling in this study to detect an inanimate reservoir; however we want to point out that on two out of four of the ICUs, sinks in most of the patient rooms were removed during the study period.Thirdly, the mutational resistome was not analysed and we only performed analysis of acquired/intrinsic resistance genes P. aeruginosa high-risk clones and carbapenemase genes, it is necessary to implement diagnostic tools at local level for their early detection and the combination with epidemiological data, in order to guide IPC strategies. This is especially the case among carbapenemase-producing high-risk clones that were associated with ongoing acquisitions. Therefore, the timely detection of carbapenemases can potentially lead to strategies to halt transmission.In conclusion, to ensure surveillance of dlac057_Supplementary_DataClick here for additional data file.


Chronic wounds that fail to heal are the most common complications experienced by diabetic patients, and current treatment remains unsatisfactory, mainly due to the vulnerability of diabetic wounds to bacterial infections. Chitosan derivatives are widely used to treat chronic wounds due to their excellent hydrophilicity, biodegradability, and antimicrobial activity and substantial contribution to tissue regeneration. However, the antimicrobial effect of chitosan is not sufficient due to the complicated pathological mechanism of diabetes mellitus. Here, we prepared carboxymethyl chitosan-grafted polyvinylpyrrolidone-iodine (CMC-g-PVPI) microspheres and used them to treat chronic wounds. Carboxymethyl chitosan (CMC) was used as the skeleton and was grafted with polyvinylpyrrolidone-iodine (PVPI) to form a CMC-g-PVPI complex hydrogel and CMC-g-PVPI microspheres, which formed as a result of the high shearing dispersion of the complex hydrogel. In vivo experiments on diabetic wounds revealed significantly accelerated wound closure in the presence of the microspheres, demonstrating the excellent potential of CMC-g-PVPI to promote skin wound regeneration under diabetic conditions.  Thus, effective wound treatment focusing on accelerated healing processes and antibacterial properties is crucial for healing diabetic wounds.Diabetes is a common global disease that threatens human lives. According to the International Diabetes Federation (IDF) Diabetes Atlas, almost half a billion people have diabetes worldwide, and this number is rapidly increasing . People 8–10]. Among the currently available biomaterials, chitosan is one of the most versatile and commercially important natural polymers. It is a cationic polysaccharide mainly derived from the deacetylation of chitin, which can be extracted from the shells of crabs and prawns [12–14]. Due to its hemostatic properties, chitosan also helps in the wound healing process. The depolymerization of chitosan releases N-acetyl-β-D-glucosamine, which induces cell proliferation and collagen deposition, further contributing to tissue regeneration . Herein, we added γ-PGA to the hydrogel to increase the viscosity and hydrophilicity of the product (CMC-g-PVPI-γ-PGA). First, we investigated the healing ability of CMC-g-PVPI-γ-PGA and found that the complex hydrogel did not present a remarkble accelerating effect. Thus, we dispersed the CMC-g-PVPI complex hydrogel into uniform microspheres using high-shear homomixers. The properties of the CMC-g-PVPI microspheres were characterized by scanning electron microscopy (SEM) and Fourier transform infrared (FTIR) spectroscopy. After testing these properties, the CMC-g-PVPI microspheres were applied to a diabetic wound surface to study their wound healing effect.In the present study, we used CMC as a skeleton and grafted PVPI onto it to form a CMC-g-PVPI complex hydrogel. The CMC-g-PVPI complex hydrogel was synthesized via state-of-the-art methodology . γ-Poly 2.2.1.CMC was purchased from Shanghai Macklin Biochemical Co., Ltd. (China). Azodiisobutyronitrile (AIBN) and methyl methacrylate were purchased from Shanghai Aladdin Biochemical Technology Co., Ltd. (China). Polyvinylpyrrolidone (PVP) was purchased from Sigma–Aldrich (Shanghai) Trading Co. Ltd. (China). γ-PGA  was purchased from Saitaisi Biological Technology Co., Ltd. (China). Iodine, potassium iodide, tert-butanol, acetic acid and sodium dodecyl sulfonate were provided by the laboratory at analytical grade.2.2.tert-butanol was added, and the solution was stirred for 1 h. Sodium dodecyl sulfonate (0.25 g) was added to the solution in the second step and stirred until it dissolved.CMC-g-PVPI products were synthesized by following a published method with some modifications . First, The white solution was added dropwise into the above 610 ml liquid through a tap funnel. The mixture was then stirred for 2 h to obtain the CMC-g-PVPI complex hydrogel. We dispersed the CMC-g-PVPI complex hydrogel into homogenized microspheres using high-shear homomixers from Shanghai Xin Sha Machinery Co., Ltd. (China), to obtain CMC-g-PVPI microspheres.Then, 400 µl water was added to 200 µl CMC-g-PVPI complex hydrogel to obtain a mixed 600 µl solution. γ-PGA (350 mg) was dissolved in the solution, and the resulting solution was ultrasonicated for 30 min in an ultrasonic cleaner. It was briefly centrifuged to produce CMC-g-PVPI-γ-PGA.2.3.Samples of CMC-g-PVPI microspheres were sputter-coated with Au and imaged by scanning electron microscopy. The acceleration voltage was 10 kV.−1, and 64 scans were performed. First, we scanned the blank background, and then we placed the sample in the instrument and measured the Fourier infrared absorption peak.FTIR spectra were obtained with a NEXUS-470 FTIR series spectrometer (Thermo Scientific Nicolet iN10) via the KBr pellet pressing method. KBr pellets and samples were ground at a ratio of 70:1 and mixed uniformly. The instrument was preheated for 20 min before the operation. Software was used to adjust the spectral range to 400–4000 cm2.4.Animals: Db/db mice were obtained from the Ninth People’s Hospital Animal Center  and the experimental protocol was approved by the Animal Care and Experiment Committee of the Ninth People’s Hospital . Fifteen diabetic mice were randomly divided into 3 groups (n = 5). An 8 mm diameter round wound was made on the center of the back of each mouse. Mice in the control group were not treated. The wounds of the mice in the gel group were treated with 200 µl hydrogel. The wounds of the mice in the microsphere group were treated with 10 mg CMC-g-PVPI microspheres. Each mouse was kept isolated in its own cage and supplied with sufficient water and food. The day the wounds were created was defined as Day 0, and photographs of the wounds were taken on Days 0, 3, 5, 7, 9, 12, and 14 to evaluate the healing status. The wound area was calculated using Image-Pro Plus microscope software by applying the following formula:2.5.The mice were all sacrificed after 14 days of treatment, and wound samples were collected and fixed in 4% paraformaldehyde overnight. Then, the samples were embedded in paraffin. Each sample was cut into 5 µm thick sections and prepared for histological staining. The formation of granulation tissue was observed by hematoxylin-eosin staining (H&E), and collagen formation was detected through Masson trichrome staining. The tissue sections were incubated with a rabbit monoclonal anti-CD31 antibody  and an anti-Ki67 antibody . An HRP-labeled polymer anti-rabbit antibody  was used as the secondary antibody. Image-Pro Plus 6 software (Media Cybernetics) was used in this study to calculate the CD31 vascularity and the dermis proliferation index.2.6.Statistical analysis was performed using GraphPad Prism Version 8.0.1 . Unpaired Student’s t test (and nonparametric tests) were used in this study. Two-tailed analyses were conducted, and P values less than 0.05 (*) and 0.01 (**) were considered significant.3.The objective of this study was to fabricate a copolymer with inherent wound healing activities and antibacterial properties to efficiently promote diabetic wound closure. We synthesized two different forms of the CMC-g-PVPI complex: the hydrogel form and the microsphere form. Then, we characterized the complexes and applied them to diabetic wounds to study their healing properties. A schematic representation of this study is shown in 3.1.The surface morphology of the CMC-g-PVPI microspheres is presented in −1, which was attributed to strong stretching vibrations of the -OH and N-H bonds. In the spectrum of the CMC-g-PVPI microsphere emulsion, the corresponding peak shifted to 3424 cm−1, and this shift may be attributed to the presence of iodine. The absorption peak at 1643 cm−1 in the spectrum of CMC can be assigned to the C = O stretching vibration, and an analogous peak appears at 1601 cm−1 in the spectrum of the CMC-g-PVPI microspheres. Overall, these analyses indicate the successful graft polymerization of CMC and PVPI.We used FITR to characterize the CMC-g-PVPI microspheres. S. aureus with or without 2% CMC-g-PVPI microspheres solution at 37°C for 24 h. Figure S1 shows that the growth of S. aureus could be inhibited by CMC-g-PVPI microspheres while there was no inhibition in the control group. Therefore, CMC-g-PVPI microspheres can prevent bacterial infections in wound healing.To evaluate the antibacterial property of CMC-g-PVPI microspheres, we cultured 3.2.To investigate and compare the healing properties of the CMC-g-PVPI products, we established a diabetic wound model and divided the mice into three groups, with 5 mice in each group. An 8 mm diameter wound was created in the back skin of each mouse. Photographs were taken at different healing times, as shown in H&E staining images are shown in Neovascularization and angiogenesis were also assessed via immunohistochemical assays. 4.Chitosan has been reported to promote the expression of TGF-β1 and the production of collagen, stimulating tissue regeneration . In FiguTo date, chitosan-based materials have been widely applied in wound healing , but mosOne of the highlighted strengths of this research is the combination of wound healing materials and bacteriostatic drugs. These two materials are both essential for wound care and are typically applied in sequential order. The complex fabricated by chemical methods integrated the healing and antibacterial effects and greatly reduced the number of procedures and the time required for patients, thus alleviating the high burden on both the family and the health care system. Another strength is the selection of chitosan as the inner skeleton. As chitosan has a long history of being used as a starting material for wound healing, its acceleration of hemostatic activities and cell proliferation provide a foundation for the healing effect. The microsphere form also plays a crucial role in the healing process, with a larger surface area to better absorb exudate.Limitations also exist in this study. We did not include a chitosan group as one of the control groups to compare the healing properties of the CMC-g-PVPI microspheres with those of CMC, which may have allowed better evaluation of the effect of the complex microspheres. Further studies could be designed to obtain more precise conclusions.5.In summary, a complex of CMC and PVPI was prepared in this study and used on the surface of diabetic wounds. It was observed that the CMC-g-PVPI microspheres had the capacity to accelerate the wound closure process. The advantage of CMC-g-PVPI microspheres is the combination of healing and antibacterial effects, resulting in a faster and novel method for wound repair.Click here for additional data file.


Multiple regression was conducted to examine the impact of these variables on problem-solving skills. The variables of religion, satisfaction with major, goal commitment, and critical thinking were found to have a significant impact on problem-solving abilities. The results were as follows: critical thinking , goal commitment , and explanatory power of 41%. To improve the nursing undergraduates’ problem-solving abilities through simulation practice, there needs to be a method that supports them in setting goals with self-leadership and enhance goal commitment. The method also needs to support the development of their critical thinking and curiosity for questions deriving from experiencing diverse programs in order to deliver effective outcomes.It has become important for nurses to implement self-leadership and exercise critical thinking in problem-solving to address the health issues of patients. This has led to a need for nursing education programs in which nursing students learn to embrace self-leadership and self-evaluation approaches to develop their skills. Within 260 nursing undergraduates with experience in simulation practice as study subjects, a self-reporting survey was conducted on self-leadership, goal commitment, critical thinking, and problem-solving skills. An analysis was conducted using the SPSS/WIN 21.0 program.  In recent nursing environments, complex and strategic practices are required, and the role of nurses is recognized for its importance. In order to effectively respond to the patients’ demands, it has become important for nurses to equip themselves with specialized skills ,2.Such change requires nurses to build not only a high level of knowledge and skills, but also self-leadership to actively solve problems with independence and autonomy, as well as critical thinking to set the best goals and to identify strategies and grounds for apply clinical judgment and decision ,4. To thInsufficient training on practice at nursing colleges leads to the decline in clinical practice capability of new nurses after their graduation. To complement this, more universities are adopting a high-performance simulator to enhance their training on practice ,9. The tDue to the characteristics of nursing training, the result of training is directly related to practice in the field, and this has led to more attention being given to the design of training courses involving skills development and training experience for nurses aiming to develop and promote their critical-thinking and problem-solving abilities ,16. HoweThe purpose of this study was to identify the relationship between self-leadership, goal commitment, critical thinking, and problem-solving abilities in nursing undergraduates who experienced simulation training on practice and the impact of these variables on problem-solving processes. This involved (1) investigating the relationship between self-leadership, goal commitment, critical thinking, and problem-solving abilities; and (2) identifying the impact of self-leadership, goal commitment, and critical thinking on problem-solving abilities.This is a descriptive investigation study conducted to identify the relationship between self-leadership, goal commitment, critical thinking, and problem-solving abilities of nursing undergraduates with the experience of simulation practice training.The participants of this study consisted of nursing undergraduates (juniors and seniors) at 4 universities who received 30 to 60 h of simulated practice training. The training was provided in one of the following two ways; one day of simulated practice training during clinical practice or five-day simulated practice training. All universities were equipped with high-performance simulators and used them in practice training. Though equipment and materials used varied by subject, high-performance simulators and dummies were used for most subjects.For the bioethics and safety of study subjects, this study was conducted after deliberation by the Institutional Review Board of University and the approval (No: KHSIRB-14-059(EA)) was obtained. During study period, the guidelines of the IRB were complied with. The description of the study was provided to study subjects and the purpose and methods of the study were explained. The survey was handed out to be filled-out by subjects who agreed to participate and they were informed the data they provided would be used solely for the study’s purpose. Study subjects were provided with a small amount of compensation in appreciation of their participation.(1)DemographicsGeneral characteristics included university, age, grade, religion, personal relationships, academic grades, major satisfaction, leadership training experience, and need for leadership. The age groups were divided into two: subjects of 22 years of age or below and subjects of 23 years of age or above. The personal relationships were classified into average, good, and very good. The major satisfaction was classified into unsatisfactory, average, satisfactory, and highly satisfactory. (2)Self-leadershipTo measure the self-leadership of nursing undergraduates, RSLQ (Revised Self-Leadership Questionnaire) developed by Houghton and Neck  was used(3)Goal commitmentTo measure the goal commitment of nursing undergraduates, the tool suggested by Klein et al.  was used(4)Critical thinking dispositionTo measure the critical thinking disposition of nursing undergraduates, the tool suggested by Yoon  was used(5)Problem-solving abilitiesThe Social Problem Solving Inventory-Revised (SPSI-R) modified by D’Zurilla and Maydeu-Olivares  and tranThe study data were collected from 18 September 2014 to 4 November 2014. The data were collected from juniors and seniors with an experience of simulated practice training from nursing departments at 4 universities in Seoul, and upon agreement for participation. The surveys were then handed out, filled-in, and retrieved straight away. A total of 280 copies were distributed and 276 copies were retrieved. With 16 surveys with insincere responses excluded, a total of 260 copies were used for final analysis.t-test and ANOVA with additional validation through Scheffe test and Fisher’s exact test. The relationship between the subjects’ self-leadership, goal commitment, critical thinking, and problem-solving abilities was analyzed using Pearson’s correlation coefficient. A multiple regression was used to identify the impact of these variables on problem-solving abilities.The data collected were analyzed using SPSS/WIN 21.0 program . The variables of mistakes, average, and percentage were used to identify general characteristics. The difference in problem-solving abilities by general characteristic was validated through The subjects of this study comprised 260 juniors and seniors in the nursing department with the experience of 30 to 60 h of simulated training on practice. The juniors accounted for 72.3% and the good personal relationships accounted for 56.5%. The major satisfaction was 55.4% and no leadership training experience was 61.9%. The results were as follows: self-leadership of 3.59 ± 0.71, goal commitment of 3.68 ± 0.57, critical thinking of 3.57 ± 0.57, and problem-solving abilities of 3.14 ± 0.58 .p = 0.003) than seniors (2.62 ± 0.46) in relation to grades. The seniors showed higher numbers of positive problem orientation  and rational problem-solving . There were significant differences in positive problem orientation  and negative problem orientation  in relation to personal relationships. The result of post-validation shows that subjects with “average (3.65 ± 0.53)” personal relationships showed a lower positive problem orientation than subjects with “very good (2.51 ± 0.72)” personal relationships. The subjects with “very good (2.51 ± 0.72)” personal relationships showed a lower negative problem orientation than subjects with “average (3.09 ± 0.66)” and “good (2.97 ± 0.71)” personal relationships. Subjects with higher academic grades showed a lower negative problem orientation , impulsive careless style , and evasive style  than subjects with lower academic grades. There was a significant difference in all sub-categories of problem-solving abilities in relation to major satisfaction. For positive problem orientation , subjects with “average (3.62 ± 0.48)” showed lower problem-solving abilities than subjects with “very satisfactory (4.10 ± 0.47)”. For negative problem orientation , subjects with “average (3.18 ± 0.68)” showed higher problem-solving abilities than subjects with “satisfactory (2.84 ± 0.69)” and “very satisfactory (2.59 ± 0.76)”. For rational problem solving , subjects with “very satisfactory (3.82 ± 0.36)” showed higher problem-solving abilities than subjects with “average (3.43 ± 0.40)” and “unsatisfactory (3.48 ± 0.69)”. For impulsive/careless style , subjects with “unsatisfactory (2.95 ± 0.50)” showed higher problem-solving abilities than subjects with “satisfactory (2.65 ± 0.51)” and “very satisfactory (2.45 ± 0.63)”. While there was a significant difference observed in relation to evasive style (F = 2.99 p = 0.019), no difference was observed in the post validation.  showed higher numbers of impulsive style , goal commitment , and self-leadership . For general characteristics, a positive correlation was observed in major satisfaction  and personal relationships  was lower than 10, showing no problem for multi-collinearity  and goal commitment (β = 0.28). Self-leadership, age, personal relationships, and leadership training experience showed no significance, with the value of inearity . Next, aNursing education and training must provide a learning environment that reflects actual practice and enables students to set up alternative measures to solve learning problems, search and obtain required data, and equip themselves with problem-solving abilities. However, the traditional top-down education systems have limitations in teaching such practical skills and are inadequate to prepare students for many challenges in the field . This stThe results of the study are similar to the previous studies; an average mean of 3.14 points for problem-solving skills, 3.36 points for problem-solving orientation, and 2.98 points for problem-solving abilities . This suThe difference in problem-solving abilities according to the general characteristics was partially significant for grades, personal relationships, and academic grades. The major satisfaction exhibited significant differences in all sub-categories. For positive problem orientation and rational problem-solving, the subjects with a greater major satisfaction showed a higher level of problem-solving skills. For negative problem orientation and impulsive/careless style, subjects with less major satisfaction showed a higher point. These results are consistent with the results of another study  on critiThe influential factors for problem-solving abilities were goal commitment and critical thinking. Although the general characteristics of age, satisfaction with nursing major, and personal relationships showed correlations with problem-solving skills, no significant result was observed in the regression analysis. In a study by Aubé et al. , a virtuAccording to the study by Han and Park , greaterTucker et al.  stated tThis study suggests that, while self-leadership showed no significant impact on problem-solving skills, it had impact on the problem-solving abilities of nursing undergraduates ,29. It wThis study holds significance in that it suggests the need for the development and implementation of various programs involving simulated practice training to enhance the problem-solving abilities of nursing undergraduates. It also suggests the need for measures to make students set their own goals and improve the level of their goal commitment, as well as the fact that the use of self-leadership may enhance the effectiveness of the process. Furthermore, it suggests the possible synergetic effects on developing critical thinking abilities by allowing students to build critical curiosity over questions they face by experiencing various programs. On the other hand, this study had limitations in that the scope of sample was limited to nursing undergraduates with the experience of simulated practice training at certain universities in Seoul city who agreed to participate in the study, suggesting that the study results should not be used for generalization. In addition, there are limitations to defining the correlation in that a cross-sectional study method was used in which the cause and outcome variables of the collected data were measured at the same time point for analysis.p < 0.01), with the explanatory power of 41%. In order to enhance the problem-solving abilities of nursing undergraduates, various education programs which involve discussions with simulated training on practice to demonstrate a step-by-step approach to problematic situations, an establishment of hypothesis, and team cooperation must be developed and implemented [This study was conducted to identify the relationship between the self-leadership, goal commitment, critical thinking, and problem-solving abilities of nursing undergraduates with the experience of simulated practice training. The multiple regression analysis results show that the variables of goal commitment and critical thinking had a significant impact on the nursing undergraduates’ problem-solving abilities (F = 23.17, lemented ,31,32. A